慢性發炎和胃癌(gastric cancer)發生及進展有關。在胃癌組織發現嗜中性球浸潤，其作用仍不清楚。腫瘤相關嗜中性球會因環境而呈現助於腫瘤生長(N1)或抑制腫瘤(N2) 的型態。以胃癌動物模式分析嗜中性球如何活化或抑制免疫反應。精胺琥珀酸合成酶 (Argininosuccinate synthetase, ASS1)將L-瓜胺酸和天門冬酸轉換形成精胺琥珀酸，再經精胺琥珀酸分解酶轉換成fumarate及精胺酸。癌化的胃上皮細胞過度表現ASS1。ASS1 和腫瘤生長或轉移以及和免疫細胞的交互作用較少被研究。本計劃的研究目的：是否抑制胃癌細胞ASS1，會抑制動物體內腫瘤生長或轉移，及研究免疫逃逸機制。本計劃的研究方式是利用所建立的新穎且獨特的小鼠原位胃癌模式，使用的胃癌細胞株-MGCC3I細胞能在小鼠的胃生長，且癌細胞會表現ASS1。利用分生技術(siRNA)，降低胃癌細胞株的ASS1的表現，這些穩定的細胞株，其ASS1的表現量較低，並和母代細胞及對照組特性比較：(1)當這些細胞株種植到小鼠的胃部或脾臟，其癌細胞的生長和轉移。(2)嗜中性球浸潤和表現細胞激素、趨化因子和黏著分子。(3)腫瘤內部嗜中性球毒殺機制。(4)腫瘤內部嗜中性球和CD8 T細胞之間聯繫。本研究計畫的重要性：(1)證明在胃癌的治療，ASS1是否是有利的免疫標的分子； (2)提供ASS1和嗜中性球以及CD8 T細胞之間交互作用的免疫調控機制。 Chronic inflammation is associated with initiation and/or progression of the most common cancer types including gastric cancer. Extensive infiltration of neutrophils was observed in gastric cancer. The role of tumor-associated neutrophils (TANs) in gastric cancer is unclear. The tumor microenvironment polarizes TANs toward a protumor (N2) versus an antitumor (N1) phenotype. It will be importance to investigate the type of neutrophils infiltrated in gastric animal tumor and how to activate/inhibit an immune response. Argininosuccinate synthetase (ASS1) catalyzes the conversion of citrulline and aspartic acid to argininosuccinate, which is then converted to arginine and fumaric acid by argininosuccinate lyase. High levels of ASS1 mRNA and protein have been noted in malignant gastric epithelium compared to corresponding normal epithelia. The role of ASS1 in gastric cancer progression/metastasis and the interaction between immune cells were less investigated. The aims of this research project are: to study whether the suppression of ASS1 in gastric cancer can repress tumor progression and/or metastasis in immune-competent host and investigate the mechanism of immune escape. The research will be performed on a novel unique mouse orthotropic gastric cancer animal model which we have established. The murine gastric carcinoma cell line-MGCC3I cells are able to grow orthotropically in mouse stomach; furthermore, ASS1 was expressed in this cell line.We will use siRNA targeting ASS1 to downregulate the endogenous expression in MGCC3I cells. The stable clones with low expression of ASS1 will be compared with parental cell, vector control and scramble RNA on the following properties: (a) growth and metastasis when implanted into stomach of immune competent mice, (b) the infiltration of neutrophils in tumor and the related cytokines, chemokines, and cell adhesion molecules in the recruitment and chemoattraction of neutrophils, (c) the mechanisms of killing by the intratumoral neutrophils, (d) to explore the intratumoral connections between TANs and CD8+ T cells. The significance of this research proposal is two-fold: (1) demonstrate whether ASS1 is a useful immunological target for gastric cancer therapy; (2) provide immune-regulation mechanism of interaction between ASS1 and neutrophil and CD8 T cell.
